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Phytoremediation of Pharmaceuticals with Salix Exigua View metadata, citation and similar papers at core.ac.uk brought to you by CORE provided by OPUS: Open Uleth Scholarship - University of Lethbridge Research Repository University of Lethbridge Research Repository OPUS http://opus.uleth.ca Theses Arts and Science, Faculty of 2006 Phytoremediation of pharmaceuticals with salix exigua Franks, Carmen G. Lethbridge, Alta. : University of Lethbridge, Faculty of Arts and Science, 2006 http://hdl.handle.net/10133/536 Downloaded from University of Lethbridge Research Repository, OPUS PHYTOREMEDIATION OF PHARMACEUTICALS WITH SALIX EXIGUA CARMEN FRANKS Bachelor of Science, University of Lethbridge, 2004 A Thesis Submitted to the School of Graduate Studies of the University of Lethbridge in Partial Fulfilment of the Requirements for the Degree MASTER OF SCIENCE Department of Biological Sciences University of Lethbridge LETHBRIDGE, ALBERTA, CANADA © Carmen G. Franks, 2006 PHYTOREMEDIATION OF PHARMACEUTICALS WITH SALIX EXIGUA CARMEN G. FRANKS Approved: September 28, 2006 Dr. Stewart B. Rood, Supervisor, Department of Biological Sciences Dr. Alice Hontela, Thesis Committee Member, Department of Biological Sciences Dr. Bryan E. Kolb, Thesis Committee Member, Department of Neuroscience - Cdn Ctr for Behavioural Neuroscience (CCBN) Dr. David M. Reid, External Examiner, University of Calgary, Department of Biological Sciences Dr. Mathew G. Letts, Chair, Thesis Examination Committee, Department of Geography ii Abstract Municipal treated wastewater entering rivers contain biologically active pharmaceuticals capable of inducing effects in aquatic life. Phytoremediation of three of these pharmaceuticals and an herbicide was investigated using Sandbar willow (Salix exigua) and Arabidopsis thaliana. Both plants were effective at removing compounds from solution, with removal of 86% of the synthetic estrogen, 17α-ethynylestradiol, 65% of the anti-hypertensive, diltiazem, 60% of the anti-convulsant, diazepam (Valium®), and 51% of the herbicide atrazine, in 24 hours. Distribution of compounds within roots and shoots, in soluble and bound forms, differed among compounds. Uptake and distribution of pharmaceuticals within the study plants confirmed pharmaceutical behaviour can be predicted based on a physiochemical property, their octanol-water partitioning coefficients. An effective method for detection of 17α-ethynylestradiol within surface water using solid phase extraction and gas chromatography-mass spectrometry was developed. Previously unreported breakdown of 17α-ethynylestradiol into another common estrogen, estrone, during preparative steps and gas chromatography was resolved. iii Preface - Thesis Structure This research-based MSc thesis includes an introductory chapter, three stand alone research chapters, and an integrative conclusion chapter. Chapter 1, ‘Introduction’, provides background information on phytoremediation and the emerging issue of pharmaceuticals entering the water environment, as well as an introduction to the contents of this thesis. Chapter 2, ‘Phytoremediation of trace pharmaceuticals, diltiazem, diazepam and 17α- ethynylestradiol with sandbar willow (Salix exigua)’, represents a stand alone research chapter. It describes the application of an abundant riparian willow species for the removal of trace pharmaceuticals from solution. The herbicide atrazine is included as a positive control, as it is already known to be taken up by plants. Chapter 3, ‘Phytoremediation of trace pharmaceuticals, diltiazem, diazepam and 17α- ethynylestradiol with Arabidopsis thaliana’, is another stand alone research chapter. The model plant Arabidopsis is investigated not for its prospective field application, but for its possible future genetic inquiry. Chapter 4, A ‘Method for detection of 17α-ethynylestradiol in surface water,’ is a methodology chapter. A method is outlined for extracting and measuring the synthetic birth control hormone ethynylestradiol from waste or surface water. iv Chapter 5, ‘Discussion,’ integrates the information from the three prior research chapters. Finally, Appendix A presents the statistical analyses conducted for the thesis, while Appendix B provides additional details of the experimental methods that are not provided in Chapters 2 or 3. Appendix C provides a figure of the nuclear magnetic resonance (NMR) analysis performed on 17α-ethynylestradiol to verify its purity. Further information on the pharmaceuticals and the herbicide used in this study are provided in Appendix D. v Acknowledgements My thanks to Stewart Rood, for providing me the tools, equipment and a corner desk within his ‘sweat shop’ to do this MSc project, and to my committee members for their support. I gratefully thank David Pearce for his brain, his seemingly endless patience and for ‘three turns widdershins’. A special thanks to David Reid for loan of the oxidizer that helped me to complete this project and Heather Bird for helping me get the oxidizer up and running. And to Shannon MacLeod, I am eternally grateful, for you helped me through this time in my life and humored me convincingly when I talked about this ‘stuff’. vi Table of Contents Signature Page .................................................................................................................... ii Abstract..............................................................................................................................iii Preface - Thesis Structure .................................................................................................. iv Acknowledgements............................................................................................................ vi Table of Contents.............................................................................................................. vii List of Tables ...................................................................................................................... x List of Figures................................................................................................................... xii Abbreviations.................................................................................................................... xv CHAPTER 1 Introduction.................................................................................................. 1 1.1 Background: Pharmaceuticals in the water environment..................................... 1 1.2 Study pharmaceuticals ......................................................................................... 4 1.3 Introduction to phytoremediation ........................................................................ 7 1.4 Study plants: Salix exigua & Arabidopsis thaliana............................................ 11 1.5 Plant physiology: root uptake and translocation................................................ 13 1.6 MSc Project summary........................................................................................ 27 Literature Cited ........................................................................................................... 30 CHAPTER 2 Phytoremediation of trace levels of pharmaceuticals, diltiazem, diazepam and 17α-ethynylestradiol, and the herbicide atrazine, with sandbar willow (Salix exigua) ........................................................................................................................................... 38 2.1 Introduction........................................................................................................ 38 2.2 Materials and Methods....................................................................................... 41 2.2.1 Chemicals..................................................................................................... 41 2.2.2 Plants, hydroponics and treatments.............................................................. 44 vii 2.2.3 Uptake studies.............................................................................................. 47 2.2.4 Root concentration factors and transpiration stream concentration factors. 50 2.2.5 Soluble fractions .......................................................................................... 53 2.2.6 Bound fractions............................................................................................ 55 2.3 Results................................................................................................................ 57 2.3.1 Uptake studies time course .......................................................................... 57 2.3.2 Distribution .................................................................................................. 63 2.3.3 Root concentration factors and transpiration stream concentration factors. 66 2.4 Discussion.......................................................................................................... 74 Literature Cited ........................................................................................................... 81 CHAPTER 3 Phytoremediation of trace levels of pharmaceuticals, diltiazem, diazepam and 17α-ethynylestradiol, and the herbicide atrazine, with Arabidopsis thaliana ........... 83 3.1 Introduction........................................................................................................ 83 3.2 Materials and Methods....................................................................................... 84 3.2.1 Chemicals..................................................................................................... 84 3.2.2 Plants, hydroponics
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